Reconstruction of metabolic pathway for isobutanol production in Escherichia coli

Abstract

Background The microbial production of useful fuels and chemicals has been widely studied. In several cases, glucose is used as the raw material, and almost all microbes adopt the Embden–Meyerhof (EM) pathway to degrade glucose into compounds of interest. Recently, the Entner–Doudoroff (ED) pathway has been gaining attention as an alternative strategy for microbial production. Results In the present study, we attempted to apply the ED pathway for isobutanol production in Escherichia coli because of the complete redox balance involved. First, we generated ED pathway-dependent isobutanol-producing E. coli . Thereafter, the inactivation of the genes concerning organic acids as the byproducts was performed to improve the carbon flux to isobutanol from glucose. Finally, the expression of the genes concerning the ED pathway was modified. Conclusions The optimized isobutanol-producing E. coli produced 15.0 g/L of isobutanol as the final titer, and the yield from glucose was 0.37 g/g (g-glucose/g-isobutanol). Electronic supplementary material The online version of this article (10.1186/s12934-019-1171-4) contains supplementary material, which is available to authorized users.

Metabolic engineering of Zymomonas mobilis for anaerobic isobutanol production

“…In addition, current efforts on improving isobutanol production in these microorganisms have been focused on increasing the pyruvate pool, eliminating by-product formation, and maintaining redox balance [44,[47][48][49][50][51]. For example, the Entner-Doudoroff (ED) pathway was applied into E. coli, and the ED pathway-dependent isobutanol-producing E. coli strain was further optimized to produce 15.0 g/L isobutanol after byproduct biosynthesis genes were inactivated [52]. Z. mobilis can be a suitable host to address these issues since it has the characteristics of a unique anaerobic ED pathway, a truncated tricarboxylic acid cycle (TCA), and simple metabolism resulting in abundant pyruvate biosynthesis with redox balancing that limits by-product formation.…”

Metabolomic analysis improves bioconversion of methanol to isobutanol in Methylorubrum extorquens AM1

“…Isobutanol extraction and quantification were conducted according to the previously established protocol. [ 35 ] Briefly, the supernatant during the cultivation was extracted with dichloromethane. The organic phase containing isobutanol was measured with GC‐MS (Shimazu QP2020) using Rtx‐5MS column (30 m × 0.25 mm × 0.25 μm film; Restek, Bellefonte).…”

“…This production was 1.5-fold higher compared with the YKB3 strain, which was improved by 370-fold compared with the initial engineered strain YKA2, while the final titer achieved by the YKB4 strain was still significantly lower than that obtained from recombinant strains of E. coli or B. subtilis. [35,47] One potential strategy for further improving the titer in the future is to increase the flux into pyruvate by reinforcing the expression of pyruvate kinase or malate enzyme. [16] Given that their substrates, PEP, and malate, are intermediates of the serine cycle, future work will be essential for fine-tuning their expressions at a proper level to avoid the risk of unbalancing the interconnected metabolic pathways of the EMC pathway and TCA cycle.…”

Nitrogen‐Neutral Amino Acids Refinery: Deamination of Amino Acids for Bio‐Alcohol and Ammonia Production

“…Protein biomass reportedly produces biogas through the anaerobic digestion of microorganisms in organic waste treatment processes 10, 17–19. Despite the development of various amino acid‐producing strains, studies on biochemical production using deaminated amino acid carbon structures by engineering amino acid metabolic pathways from sugar‐based biomass such as glucose and xylose are scarce 20–23. Therefore, in this review paper, we present a case study on biochemical production through the enzyme‐based deamination reaction of amino acids in cells.…”